Spinal cord injury (SCI) is a long-term health care problem in the United States, and with the exception of the modestly effective methylprednisolone, there is currently no neuroprotective intervention clinically available for treatment of acute SCI. Our published data and preliminary results demonstrate that oxidative damage to key mitochondrial enzymes and subsequent mitochondrial dysfunction is key to the neuropathological sequalae following SCI. This proposal focuses on directly targeting mitochondrial dysfunction as a novel therapeutic intervention for contusion SCI, the fundamental concept being that SCI-induced excitotoxicity increases mitochondrial Ca2+ cycling/overload and the production of reactive oxygen species (ROS), ultimately leading to mitochondrial dysfunction and glutathione (GSH) depletion. Our approach is two-pronged, aimed at reducing mitochondrial ROS production utilizing a novel, cell-permeant antioxidant and GSH precursor, NACA (the amide form of N-acetylcysteine), as well as an alternative biofuel substrate for energy production, acetyl-l-carnitine (ALC), following SCI. Our published and preliminary data signify that both NACA and ALC improve mitochondrial bioenergetics following contusion SCI in rats, and that prolonged NACA or ALC treatment increases tissue sparing following injury. The planned experiments are designed to test the novel hypothesis that reducing oxidative damage to key mitochondrial proteins will maintain mitochondrial bioenergetics, thus leading to increased neuroprotection and improved functional recovery following contusion SCI. Specifically we will: 1) Characterize oxidative damage to specific mitochondrial proteins involved in bioenergetics and test the hypothesis that NACA treatment ameliorates mitochondrial oxidative damage following SCI, 2) Test the hypothesis that a combinatorial treatment with NACA and ALC will act synergistically to preserve mitochondrial homeostasis following SCI, and 3) Test the hypothesis that a combinatorial treatment with NACA and ALC will increase tissue sparing and promote long-term functional recovery following SCI. Critically, this application is built around the utilization of several novel techniques we have developed for isolating synaptic (neuronal) and non-synaptic (soma and glia) mitochondria from the injured spinal cord, as well as an L1/L2 contusion SCI paradigm that demonstrates a significant correlation between neuroprotection and remarkable improvements in recovery of hind limb function. Collectively, the proposed experiments will pinpoint key mitochondrial events that could be potential novel targets for pharmacological interventions to more effectively treat SCI and, perhaps, other CNS injuries.